Within the scope of the present invention, a microparticle or a microcarrier refer to any type of particles, respectively to any type of carriers, microscopic in size, typically with the largest dimension being from 100 nm to 300 micrometers, preferably from 1 μm to 200 μm.
According to the present invention, the term microcarrier refers to a microparticle functionalized, or designed to be functionalized, that is containing, or designed to contain, one or more ligands or functional units bound to the surface of the microcarriers or impregnated in its bulk. A large spectrum of chemical and biological molecules may be attached as ligands to a microcarrier. A microcarrier can have multiple functions and/or ligands. As used herein, the term functional unit is meant to define any species that modifies, attaches to, appends from, coats or is covalently or non-covalently bound to the surface of said microcarrier or impregnated in its bulk. These functions include all functions that are routinely used in high-throughput screening technology and diagnostics.
Any reference in this disclosure to a code of a microcarrier or of a microparticle includes codes written on the surface of said microcarrier, or of said microparticle, as well as codes written at an internal depth of the microcarrier or microparticle. Such codes and methods for writing codes are disclosed, for example, in the patent application WO 00/63695 which is herein incorporated by reference. In particular, all aspects of the patent application WO 00/63695 related to the codes and the methods for writing and reading are herein specifically incorporated by reference.
Drug discovery or screening and DNA sequencing commonly involve performing assays on very large numbers of compounds or molecules. These assays typically include, for instance, screening chemical libraries for compounds of interest or particular target molecules, or testing for chemical and biological interactions of interest between molecules. Those assays often require carrying out thousands of individual chemical and/or biological reactions.
A number of practical problems arise from the handling of such a large number of individual reactions. The most significant problem is probably the necessity to label and track each individual reaction.
One conventional method of tracking the identity of the reactions is achieved by physically separating each reaction in a microtiter plate. The use of microtiter plate, however, carries several disadvantages like, in particular, a physical limitation to the size of microtiter plate used, and thus to the number of different reactions that may be carried out on the plate.
In light of the limitations in the use of microarray, they are nowadays advantageously replaced by functionalized encoded microparticles to perform chemical and/or biological assays. Each functionalized encoded microparticle is provided with a code that uniquely identifies the particular I′ ligand(s) bound to its surface. The use of such functionalized encoded microparticles allows for random processing, which means that thousands of uniquely functionalized encoded microparticles may all be mixed and subjected to an assay simultaneously. Examples of functionalized encoded microparticles are described in the international patent application WO 00/63695 and are illustrated in FIG. 1.
The international patent application WO 2010/072011 describes an assay device having at least a microfluidic channel which serves as a reaction chamber in which a plurality of functionalized encoded microparticles 1 (FIG. 1) can be packed. The microfluidic channel is provided with stopping means acting as filters that allow a liquid solution containing chemical and/or biological reagents to flow through while blocking the functionalized encoded microparticles 1 inside. The geometrical height of said microfluidic channels and the dimensions of said functionalized encoded microparticles 1 are chosen so that said microparticles are typically arranged in a monolayer arrangement inside each microfluidic channels preventing said microparticles 1 to overlap each other.
Those functionalized encoded microparticles 1 that show a favorable reaction of interest between their attached ligand(s) and the chemical and/or biological reagents flowing through may then have their code read, thereby leading to the identity of the ligand that produced the favorable reaction.
The term microfluidic channel refers to a closed channel, i.e. an elongated passage for fluids, with a cross-section microscopic in size, i.e. with the largest dimension of the cross-section being typically from about 1 to about 500 micrometers, preferably about 10 to about 300 micrometers. A microfluidic channel has a longitudinal direction, that is not necessarily a straight line, and that corresponds to the direction in which fluids are directed within the microfluidic channel, i.e. preferably essentially to the direction corresponding to the average speed vector of the fluid, assuming a laminar flow regime.
With the assay device described in WO 2010/072011, the detection of a reaction of interest can be based on continuous readout of the fluorescence intensity of each functionalized encoded microparticle 1 present in a microfluidic channel, as depicted in FIG. 6a. FIG. 6a clearly shows that it is difficult or even impossible to extract early quantitative information from the slopes at the origin when considering the intensity of each functionalized encoded microparticle 1 as a function of time. Therefore, the functionalized encoded microparticles 1 and the assay device described in WO 2010/072011 do not allow for a rapid quantification of reagent or ligand before an equilibrium state is reached, when the fluorescent signals saturate. Although the assay device of WO 2010/072011 decreases the time needed to reach equilibrium, in typical concentration values of analyte in the nano-molar range, ten to twenty minutes are still required, while lower concentration in the pico-molar range can take up to hours to be reached and serve for quantification. Moreover, the discrepancies in their fluorescent signals, in particular the diffusion pattern even after the equilibrium has been reached does not determine a quantitative information with a lower margin of error than about 15%.
The present invention aims to remedy all or part of the disadvantages mentioned above.